Test plates for chemical or biochemical analyses, which contain a plurality of individual wells or reaction chambers, are well-known laboratory tools. Such devices have been employed for a broad variety of purposes and assays, and are illustrated in U.S. Pat. Nos. 4,734,192 and 5,009,780, for example. Microporous membrane filters and filtration devices containing the same have become particularly useful with many of the recently developed cell and tissue culture techniques and assays, especially in the fields of bacteriology and immunology. Multiwell plates, used in assays, often utilize a vacuum applied to the underside of the membrane as the driving force to generate fluid flow through the membrane.
The microplate has been used as a convenient format for plate processing such as pipetting, washing, shaking, detecting, storing, etc. A variety of assays have been successfully formatted using multiwell filter plates with vacuum driven follow-through. Applications range from Cell Based assays, genomics and proteomic sample prep to immuno-assays.
An example of a protein digestion sample process may include the following steps:
1. Deposit the protein sample in the wells with the digestion enzymes.
2. Bind or capture the digested protein in or on the filter structure.
3. A series of sample washes where the solutions are transferred to waste by vacuum.
4. Solvent elution to recover the concentrated sample.
Another filter plate application used for a Genomic Sequencing Reaction Clean-up may include the following steps:
1. Deposit the sample into the wells and concentrate product onto the membrane surface by vacuum filtration to waste.
2. A series of sample washes where the solutions are transferred to waste by vacuum. Repeated and then filter to dryness.
3. Re-suspend the sample on the membrane and aspirate off the re-suspended sample from the membrane surface.
Washing to waste is easily accomplished with virtually any of the conventional manifolds available. During a wash step, a relatively large volume (greater than 50 .mu.l) of aqueous solution is added to the wells and drawn to waste. The orientation of the plate is not critical when adding a large volume of liquid, as long as the transfer pipette or other device is able to access the well opening. However with the Protein Digestion example, the elution volumes are relatively small (less than 15 .mu.l) and can be as low as about 1 .mu.l. This small volume needs to be deposited directly on the filter structure in the well to insure the solvent is drawn through the structure for complete elution of the sample. With the other example, Sequencing Reaction Clean-up, the final concentrated sample is between 10-20 .mu.l and must be aspirated off the membrane without damaging the membrane surface.
Many of these and other protocols require the addition of small accurate liquid volumes. When using filter bottom plates the performance benefit is achieved because of the follow-through nature of the filter. To achieve flow through the filter a pressure differential is applied. When using automated equipment, vacuum filtration is the preferred method because of its convenience and safety. To filter by vacuum, many manufacturers provide a vacuum manifold for their products and equipment. Still, accurate liquid transfer is not possible on the deck of a conventional liquid handler, because the position of the plate in the Z-direction can vary during use. Indeed, all of the standard manifolds available today use a compressible gasket material to seal the filter plate, and during the evacuation of the vacuum chamber in the manifold, the plate moves as the gasket is compressed. The amount of plate movement varies, depending in part upon the durometer of the gasket used and the vacuum pressure that is applied. The amount of movement is too great or variable to be able to program a liquid handling robot to account for the movement, making successful, reproducible automated transfer difficult or impossible. Similar problems arise with the Sequencing clean-up where the small volume is aspirated off the surface of the membrane. If the position of the membrane varies then it is not possible to program the automated equipment to aspirate off the surface of the membrane without potentially damaging the membrane surface.
Additionally, to insure quantitative transfer of filtrate from a 384-well filter plate into a collection plate, the spouts must be as close to the collection plate openings as possible. The available manifolds have a gasket sealing to the underside of the filter plate, and thus the only way to use these manifolds to achieve quality transfers is to have the spouts extend below the plate flange and into the wells of the collection plate. However, in such a design, the spouts are exposed and are thus prone to damage and/or contamination.
It is therefore an object of the present invention to provide a vacuum manifold assembly that is readily adapted to automation protocols.
It is another object of the present invention to provide a vacuum manifold assembly that fixes the position of a sample-processing device, such as a multiwell plate, regardless of the vacuum applied.
It is a further object of the present invention to provide a vacuum manifold assembly with features that enable quantitative filtrate transfer to a collection well when used with multiwell plates with dense arrays of wells.
It is another object of the present invention to provide a vacuum manifold assembly that enables direct transfer on an analytical device such as and MALDI target.
It is still another object of the present invention to provide a vacuum manifold assembly that is modular and adaptable to a variety of applications.